Project Summary/Abstract PALB2 is tumor suppressor protein that physically and functionally links BRCA1 and BRCA2, the two major breast cancer suppressors, in the DNA damage response and tumor suppression. The 3 proteins function in a BRCA1-PALB2-BRCA2 axis to play essential roles in in homologous recombination (HR)-based DNA double strand break repair (DSBR) and cell cycle checkpoint control after DNA damage. These functions are critical for the maintenance of genome stability and suppressing tumorigenesis. In recent years, much has been learned about the mechanisms of PALB2 in HR-DSBR, but the mechanisms by which PALB2 functions with BRCA2 and BRCA1 to promote cell cycle checkpoint response remain elusive, and how normal PALB2 mutant cells survive and proliferate in vivo and evolve into cancer cells is poorly understood. In this proposal, we will deploy a combination of state-of-the-art in vitro and in vivo approaches to answer the above open questions. In Aim 1, we will use phosphoproteomics, targeted proteomics and functional studies to identify key factors and mechanisms of PALB2-mediated G2/M checkpoint control. In Aim 2, we will build upon our recent discovery of NF?B activation in Palb2 mutant mice and use a combination of systemic knockin, conditional knockout, lineage tracing and organoid models to define the mechanisms of PALB2-associated mammary tumorigenesis and the role of NF?B in the process. In Aim 3, we will determine the genetic mechanisms of our newly discovered complementary relationship of PALB2 and BRCA1 in mammary tumor development by testing the roles of RAD52 and RNF168 in the respective models.